The present invention relates to techniques for improving the activity of an immobilized lipase and for regenerating a deactivated immobilized lipase, wherein said immobilized lipase is particularly useful in a method of generating biodiesel by transesterification of triglycerides and a lower alcohol.
In many countries, e.g. Europe, U.S.A., and Japan, etc., strong bases have been used as a catalyst in industrial processes for producing biodiesel, for example U.S. Pat. No. 5,354,878. Such industrial processes have a production scale of hundreds of thousands of tons. Even so, a strong base process suffers some serious defects. For example, a strong base process can not smoothly handle rendered oils and fats which contain a higher content of impurities. The impurities described herein mainly are moisture and free fatty acids commonly contained in rendered feedstock oils, rendered animal fats, and rendered oils and rendered fats generated in the refining of feedstock oils. The existence of these impurities will cause a strong base process to generate many undesirable by-products (e.g. soap), thereby lowering the yield of biodiesel and making purification of the biodiesel product more difficult. Therefore, nearly all commercial strong base processes for producing biodiesel use pure vegetable oils as the raw material.
At present, there are two major key difficulties in using lipase to produce biodiesel. The first difficulty is that the activity of lipase is relatively low. In an article presented by Watanabe, et al., [xe2x80x9cContinuous Production of Biodiesel Fuel from Vegetable Oil Using Immobilized Candida antarctica Lipasexe2x80x9d, JAOCS, vol. 77, pp. 355-360, 2000], the lipase process requires 36 hours to complete the reaction which is significantly longer than one hour required by the strong base process. Another difficulty is that the price of lipase is much higher than the price of sodium hydroxide (NaOH) used by the strong base process. Unless an immobilized lipase having an enhanced activity is used, and the activity of the immobilized lipase can be maintained after being recycled for a certain number of times, a lipase process is difficult to compete with the strong base process in terms of the production cost. Unfortunately, the immobilized lipase is liable to be poisoned by a lower alcohol, and the deactivated immobilized lipase can not be regenerated effectively with its recovered activity being comparable to that of the parent immobilized lipase. Therefore, in order to make an immobilized lipase process becoming economically feasible, or even replacing the conventional strong base process, issues such as how to increase the activity and life-span of an immobilized lipase, and how to effectively regenerate an immobilized lipase that is partially or completely deactivated, have become very important.
One objective of the present invention is to provide a method suitable for enhancing the activity of an immobilized lipase.
Another objective of the present invention is to provide a method for regenerating an immobilized lipase having a reduced activity.
Still another objective of the present invention is to provide a method of preparing a lower alkyl fatty acid ester, in particular biodiesel, by transesterification or esterification of a fatty acid glyceride or a free fatty acid with a lower alcohol using a pretreated or regenerated immobilized lipase as the catalyst.
The inventors of the present invention deem that a decrease in the activity of transesterification of a fatty acid glyceride and a lower alcohol is mainly caused by physical factors, i.e. the immiscibility between methanol or ethanol and fatty acid glycerides. Consequently, when methanol or ethanol is absorbed into the voids of an immobilized lipase, the entry of fatty acid glyceride into the voids will be blocked, stopping the reaction from taking place. The inventors also observe that methanol is easier to be absorbed by the immobilized lipase than an oil. The inventors of the present invention first disclose an ideal solvent to wash a deactivated immobilized lipase. This solvent needs to be harmless to the lipase, and has a good solubility to oil, grease, moisture, and methanol or ethanol. For example, an alcohol with three or more than three carbon atoms, preferably iso-propanol, 2-butanol and tert-butanol, can effectively regenerate a deactivated immobilized lipase. The inventors also discover that the activity of an immobilized lipase can be significantly increased when such an ideal solvent is used to perform an immersion pretreatment on an immobilized lipase.
In a preferred embodiment of the present invention, the activity of a commercially available immobilized lipase Novozyme 435 was increased 8-10 times in comparison with the one receiving no pretreatment; and a deactivated Novozyme 435 was successfully regenerated to an activity level equivalent to the level before being poisoned.